Inhibitory Factors Research Articles

Diversity of Memory CD8+ T Cells in Tumor-Draining Lymph Nodes from Patients with Bladder Cancer.

The role of memory T cells in orchestrating memory responses to previously known tumor antigens is well documented. The aim of this study was to assess the frequency of different memory T cell subsets in tumor-draining lymph nodes of patients with bladder cancer (BC) and their prognostic significance. Mononuclear cells were isolated from 50 tumor-draining lymph nodes of untreated patients with BC and stained with antibodies against the markers CD8, CD95, CD45RO and CCR7. Data were collected using the FACSCalibur flow cytometer and analyzed using FlowJo software. Among the CD8+ cytotoxic lymphocytes, the frequency of different subsets was determined including total memory cells (CD8+CD45RO+CD95+), T central memory (TCM: CD8+CCR7+CD45RO+CD95+), T effector memory (TEM: CD8+CCR7-CD45RO+CD95+), T stem cell memory (TSCM: CD8+CCR7+CD45RO-CD95+) and naïve T cells (CD8+CCR7+CD45RO-CD95-). The analysis revealed that on average 49.32±20.15 (between 1.62% and 87.20%) percent of CD8+ lymphocytes in draining lymph nodes of BC had a memory phenotype. TCM cells showed the highest frequency (34.71±17.04), while TSCM cells (7.51±8.53) demonstrated the lowest. The total frequency of memory cells tended to be higher in patients with tumor invasion to muscle layer (P=0.052) and stage III (P=0.042) than in patients without invasion and stage I. The TCM subset was more frequent in patients with necrotic tumors than in patients without necrosis (P=0.048). TSCM significantly increased in patients with N2 compared to N0 (P=0.042). Conversely, the ratio of TSCM cells to total memory cells was higher in lower tumor stages (P=0.059), tumors without muscle invasion (P=0.026) and low T grouping (P=0.043). Overall the data indicated an increase in the frequency of memory T cells and their TSCM and TCM cells with tumor progression. In contrast, the ratio of TSCM to total memory cells was higher in less advanced tumors. These results suggest that the immune system is frequently exposed to tumor antigens and strives to create a memory T cell reservoir, but this is suppressed by inhibitory factors provided by the tumor. These findings emphasize the importance of understanding the dynamic interplay between memory T cell subsets and BC progression.

Comprehensive Analysis of the Immunosuppressive Function of Regulatory T Cells in Human Hepatocellular Carcinoma Tissues.

Immune-based therapies are commonly employed to combat hepatocellular carcinoma (HCC). However, the presence of immune-regulating elements, especially regulatory T cells (Tregs), can dramatically impact the treatment efficacy. A deeper examination of the immune-regulation mechanisms linked to these inhibitory factors and their impact on HCC patient outcomes is warranted. We employed multicolor fluorescence immunohistochemistry (mIHC) to stain Foxp3, cytokeratin, and nuclei on an HCC tissue microarray (TMA). Leveraging liver cancer transcriptome data from TCGA, we built a prognostic model focused on Treg-associated gene sets and represented it with a nomogram. We then sourced liver cancer single-cell RNA sequencing data (GSE140228) from the GEO database, selectively focusing on Treg subsets, and conducted further analyses, including cell-to-cell communication and pseudo-time trajectory examination. Our mIHC results revealed a more substantial presence of Foxp3+Tregs in HCC samples than in adjacent normal tissue samples (P < .001). An increased presence of Foxp3+Tregs in HCC samples correlated with unfavorable patient outcomes (HR = 1.722, 95% CI:1.023-2.899, P = .041). The multi-factorial prognosis model we built from TCGA liver cancer data highlighted Tregs as a standalone risk determinant for predicting outcomes (HR = 3.84, 95% CI:2.52-5.83, P < .001). Re-analyzing the scRNA-seq dataset (GSE140228) showcased distinctive gene expression patterns in Tregs from varying tissues. Interactions between Tregs and other CD4+T cell types were predominantly governed by the CXCL13/CXCR3 signaling pathway. Communication pathways between Tregs and macrophages primarily involved MIF-CD74/CXCR4, LGALS9/CD45, and PTPRC/MRC1. Additionally, macrophages could influence Tregs via HLA-class II and CD4 interactions. An elevated presence of Tregs in HCC samples correlated with negative patient outcomes. Elucidating the interplay between Tregs and other immune cells in HCC could provide insights into the modulatory role of Tregs within HCC tissues.

Distinct characteristics of unique immunoregulatory canine non-conventional TCRαβpos CD4negCD8αneg double-negative T cell subpopulations.

Conventional CD4pos regulatory T (Treg) cells characterized by expression of the key transcription factor forkhead box P3 (FoxP3) are crucial to control immune responses, thereby maintaining homeostasis and self-tolerance. Within the substantial population of non-conventional T cell receptor (TCR)αβpos CD4negCD8αneg double-negative (dn) T cells of dogs, a novel FoxP3pos Treg-like subset was described that, similar to conventional CD4pos Treg cells, is characterized by high expression of CD25. Noteworthy, human and murine TCRαβpos regulatory dn T cells lack FoxP3. Immunosuppressive capacity of canine dn T cells was hypothesized based on expression of inhibitory molecules (interleukin (IL)-10, cytotoxic T-lymphocyte associated protein 4, CTLA4). Here, to verify their regulatory function, the dnCD25pos (enriched for FoxP3pos Treg-like cells) and the dnCD25neg fraction, were isolated by fluorescence-activated cell sorting from peripheral blood mononuclear cells (PBMC) of Beagle dogs and analyzed in an in vitro suppression assay in comparison to conventional CD4posCD25pos Treg cells (positive control) and CD4posCD25neg T cells (negative control). Canine dnCD25pos T cells suppressed the Concanavalin A-driven proliferation of responder PBMC to a similar extent as conventional CD4posCD25pos Treg cells. Albeit to a lesser extent than FoxP3-enriched dn and CD4posCD25pos populations, even dnCD25neg T cells reduced the proliferation of responder cells. This is remarkable, as dnCD25neg T cells have a FoxP3neg phenotype comparable to non-suppressive CD4posCD25neg T cells. Both, CD25pos and CD25neg dn T cells, can mediate suppression independent of cell-cell contact and do not require additional signals from CD4posCD25neg T cells to secrete inhibitory factors in contrast to CD4posCD25pos T cells. Neutralization of IL-10 completely abrogated the suppression by dnCD25pos and CD4posCD25pos Treg cells in a Transwell™ system, while it only partially reduced suppression by dnCD25neg T cells. Taken together, unique canine non-conventional dnCD25pos FoxP3pos Treg-like cells are potent suppressor cells in vitro. Moreover, inhibition of proliferation of responder T cells by the dnCD25neg fraction indicates suppressive function of a subset of dn T cells even in the absence of FoxP3. The identification of unique immunoregulatory non-conventional dn T cell subpopulations of the dog in vitro is of high relevance, given the immunotherapeutic potential of manipulating regulatory T cell responses in vivo.

Psychometric properties of the Mexican version of the Intolerance of Uncertainty Scale: The IUS-12M

Background: The Intolerance of Uncertainty Scale short version (IUS-12) has proven to be a robust self-report measure to assess intolerance of uncertainty. While previous psychometric analyses of the IUS-12 have established a stable two-factor structure corresponding to the prospective and inhibitory factors of intolerance of uncertainty, recent studies suggest that the bifactor model may better explain its factor structure. Objective: The current study aimed to adapt and validate the IUS-12 in a Mexican population culturally. Method: The study consisted of a convenience non-probabilistic community sample of 405 adults between 18 and 70 years. Results: Confirmatory factor analyses supported a bifactor model and good internal consistency. Invariance testing indicated partial invariance across women and men. Convergent validity tests showed that the IUS-12 was related to measures of worry, as well as depression and anxiety. Conclusion: These findings provide evidence for the reliability and validity of the adapted version of the IUS-12 in Mexico.

Effectiveness and Outcomes of Fishing Gear Assistance on Fishermen's Income in Pedes District Karawang

Fishing gear assistance is one of a series of activities undertaken by the Ministry of Marine Affairs and Fisheries by providing fishing gear, Gill Net and Trammel Net to fishermen. The study aims to analyse the level of effectiveness of activities, analyze the strength of supportive and inhibitory factors that influence, and analyse the outcome of fishing gear assistance on fishermen's reception. The research method used is quantitative descriptive. Primary data collection is obtained directly through questionnaires and interviews whereas secondary data is acquired through journal literacy and related scripts as well as data from related agencies. Sampling was done with a census of 40 fishermen. Data analysis on this study uses the likert scale, the guttman scale and the t test. The results of this study show that the overall distribution of aid is running effectively. Both supportive factors have strong influence and both inhibitory factors have weak influence in the distribution of aid. There has been a change in fishermen's reception before and after the distribution of fishing gear with a sig. (2-tailed) value of 0,000 &lt; 0,05, whereas fisherman recipient of shrimp net assistance has not experienced any change in reception after the assistance of fish gear.

Pelayanan PT. PLN (PERSERO) Unit Layanan Pelanggan (ULP) Panam Dalam Menanggapi Keluhan Pelanggan

Indonesia is one of the developing countries, to carry out fair and equitable development throughout Indonesia, there is a need for sufficient energy as a driving motor for development itself. Electricity distribution in Pekanbaru, as in other regions of Indonesia, is handled by the State Electricity Company (PLN) as the largest electricity supply company in Indonesia. PT. PLN (Persero) Pekanbaru ULP Panam branch as one of SOEs that aims to provide services in the field of electricity to the community (Public Utility). Organizational structure that is a primary and absolute requirement for increasing employment, responsibility and achievement of established goals. The purpose of this study is to describe and analyze PT.PLN services in handling customer complaints and identify and analyze the inhibitory factors of PT.PLN services in handling customer complaints. This study used Zeithaml's theory of five indicators: Tangible, Reliability, Responsiveness, Assurance, Emphathy. The methods used in this study are qualitative research methods that are descriptive. With data collection techniques, observation interviews and documentation. The service provided by PLN ULP Panam is still insufficient because of the many customer complaints that resulted in the service process being hampered. The service provided by the officers is not sufficient in responding to customer complaints and the SOP or Service Standards for officers in handling customer complaints do not exist. PLN ULP Panam must take this problem seriously because if there is no SOP or Service Standard, it is detrimental to the service process provided to customers.

Oncolytic viruses improve cancer immunotherapy by reprogramming solid tumor microenvironment.

Immunotherapies using immune checkpoint inhibitors (ICIs) and chimeric antigen receptor (CAR) T-cell therapy have achieved successful results against several types of human tumors, particularly hematological malignancies. However, their clinical results for the treatment of solid tumors remain poor and unsatisfactory. The immunosuppressive tumor microenvironment (TME) plays an important role by interfering with intratumoral T-cell infiltration, promoting effector T-cell exhaustion, upregulating inhibitory molecules, inducing hypoxia, and so on. Oncolytic viruses are an encouraging biocarrier that could be used in both natural and genetically engineered platforms to induce oncolysis in a targeted manner. Oncolytic virotherapy (OV) contributes to the reprogramming of the TME, thus synergizing the functional effects of current ICIs and CAR T-cell therapy to overcome resistant barriers in solid tumors. Here, we summarize the TME-related inhibitory factors affecting the therapeutic outcomes of ICIs and CAR T cells and discuss the potential of OV-based approaches to alleviate these barriers and improve future therapies for advanced solid tumors.

Sociodemographic factors affecting not receiving COVID-19 vaccine in Japan among people who originally intended to vaccinate: a prospective cohort study.

Vaccine hesitancy is a major issue for acquiring herd immunity. However, some individuals may go unvaccinated owing to inhibitory factors other than vaccine hesitancy. If there is even a small number of such people, support is needed for equitable vaccine distribution and acquiring herd immunity. We investigated sociodemographic factors that affected not undergoing COVID-19 vaccination in Japan among individuals who had strong intention to vaccinate before beginning the vaccination. We conducted this prospective cohort study on workers aged 20-65 years from December 2020 (baseline), to December 2021 using a self-administered questionnaire survey. There were 27,036 participants at baseline and 18,560 at follow-up. We included 6,955 participants who answered yes to this question at baseline: "Would you like to receive a COVID-19 vaccine as soon as it becomes available?" We applied multilevel logistic regression analyses to examine the association between sociodemographic factors and being unvaccinated at follow-up. In all, 289 participants (4.2%) went unvaccinated. The odds ratios (ORs) for being unvaccinated were significantly higher for participants aged 30-39 and 40-49 than those aged 60-65 years. Being divorced, widowed, or single, having low income, and having COVID-19 infection experience also had higher ORs. We found that some participants who initially had strong intention to vaccinate may have gone unvaccinated owing to vaccine side effects and the financial impact of absenteeism due to side effects. It is necessary to provide information repeatedly about the need for vaccination as well as social support to ensure that those who intend to vaccinate are able to do so when aiming for acquiring herd immunity through vaccination against COVID-19 as well as other potential infection pandemics in the future.

ПОДДЕРЖКА АДАПТАЦИИ ИНОСТРАННЫХ СТУДЕНТОВ С ВЫСШИМ ОБРАЗОВАНИЕМ: ИНТЕГРАЦИЯ ФОРМАЛЬНОГО И НЕФОРМАЛЬНОГО ОБРАЗОВАНИЯ

The article analyzes the main barriers and ways to support the adaptation of foreign students with higher education. These are graduates of foreign universities who are preparing for master's or postgraduate studies in the Russian Federation. The author details the factors that accelerate or slow down the adaptation processes of the selected student population. The main inhibitory factors are the age of students and the need to prepare for mastering educational material of particular complexity. At the same time, the experience of receiving previous higher education at home mitigates the difficulties of academic adaptation. The basis of activities to support the adaptation of foreign students at Peter the Great St. Petersburg Polytechnic University is the developed and implemented innovative model based on the principles of self-control and self-development. Volunteer initiatives occupy an important place in this model. The educational components of volunteering consist primarily of student participation in non-formal education programs, as well as mentoring, which is typical for informal education. Another important aspect of supporting academic adaptation at the postgraduate stage is the preparation of foreign students for research work in the context of project-team activities in a Russian-speaking environment. The author provides statistics that demonstrate the effectiveness of the proposed adaptation support model. In traditional models of adaptation support, a foreign student acts as a relative passive participant in adaptation activities prepared by the university. The proposed model makes a foreigner not only an active participant in adaptation processes, but also it contributes to the formation of such qualities that allow student to become an adapter for new generations of students.

Circulating Exosomal miRNA Signature As a Potential Risk Factor for DNA Virus Infection in Patients with Multiple Myeloma

Introduction DNA virus infection is a significant cause of morbidity and mortality in patients with multiple myeloma (MM). Monocyte dysfunction in MM patients plays a central role in infectious complications, but the precise molecular mechanism underlying the reduced monocyte resistance to viruses and risk factors for infection in newly diagnosed MM patients remains to be elucidated. Methods To determine the role of exosomal miRNAs in DNA infection of MM patients, we analyzed miRNAs in BM exosomes from MM patients (Exo-miRs) using high-throughput small RNA sequencing. To further evaluate the effect of these five upregulated Exo-miRs in antiviral response, we transfected the host cells with chemically synthesized miRNAs mimics to overexpress miRNAs. We utilized qPCR and WB to characterize the fate the cGAS-STING signaling pathway of THP-1 after overexpress miRNAs. The efficacy and safety of these Exo-miRs in innate antiviral immunity were evaluated using 5TGM1 C57BL/KaLwRij mice models. Additionally, miRNAs copy number as well as the viral susceptibility of newly diagnosed MM patients were determined in 82 samples from newly diagnosed biopsy proven MM patients. Results Screening of miRNAs enriched in exosomes delivered by primary MM cells identified five miRNAs as inhibitory factors that negatively regulate the cGAS-STING antiviral immune response. When engraftments of 5TGM1-Luc cells via tail vein for eight weeks in C57BL/KaLwRij mice resulted in MM progression with typical MM phenotypes, model mice were primed with Exo-miR antagomiRs oligonucleotides mixture through tail vein injection three times every 24 h followed by Murine cytomegalovirus (MCMV) challenge for 36 h. Consistent with the immunosuppression in MM patients, 5TGM1 C57BL/KaLwRij mice showed lower IFN-β in sera. Notably, intravenous administration of antagomiRs against miRNAs resulted in a marked increase of IFN-β levels in serum and IFN-I expression in organs (Figure 1). To investigate the potential of the miR-score as a prognostic determinant of vulnerability to DNA virus infection, we compared the miR-score in plasma exosomes collected from 82 newly diagnosed biopsy proven MM patients. We classified the viral susceptibility of newly diagnosed MM patients into two groups: I, DNA virus infection; II, no DNA virus infection. In human, a pronounced inverse correlation between IFN-β production by DNA virus in PBMCs and the microRNA (miR)-score of the exosomes within the plasma. The patients with DNA virus infection showed significantly enhanced circulatory expression of miRNAs than without DNA virus infection (p&amp;lt;0.001) patients. The diagnostic potential of miRNAs was assessed by plotting ROC curve, 5 miRNAs showed optimum sensitivity and specificity（AUC 0.854, P&amp;lt;0.0001, Sensitivity 87.5%, Specificity 74.1%, Cut-off values 5.234)（Figure 2）. Conclusions These findings identify a mechanism by which MM cells can suppress the host's innate immunity and potentially lead to immunocompromise in MM patients. Furthermore, the aberrant expression of specific miRNAs in MM patients emerges as a potential therapeutic target in MM cells. By quantifying miRNA scores based on abundant exosomal miRNAs in peripheral blood, the risk of DNA virus infection in MM patients can be assessed. Our findings shed light on a previously unknown molecular mechanism underlying immunosuppression in MM patients who have not received antineoplastic therapy, and also reveal a distinct miRNA signature that serves as a crucial determinant of the susceptibility of newly diagnosed MM patients to DNA virus infections.

Two Players, One Goal: BiTE ® Vs CART Targeting FLT3 in AML

Translation of the success of bispecific T cell engagers (BiTE®) and CAR T cells from B-cell to myeloid malignancies has been challenging. Identifying suitable target antigens in myeloid malignancies has been hampered by expected on-target-off leukemia toxicity. FLT3 presents with a rather favorable expression profile due to broad AML expression and limited expression within the normal hematopoietic compartment (Brauchle et al., 2020). Importantly, expression on AML bulk cells and leukemic stem cells is observed independent of FLT3 mutational status. Here, we compared two T-cell based immunotherapy approaches, BiTE® vs CART, in targeting FLT3 in AML. We tested BiTE® and CAR-mediated cytotoxicity against several AML cell lines and primary AML (pAML) cells in vitro. On-target off-leukemia toxicity was evaluated in mixing assays of pAML cells and healthy donor bone marrow (hBM). The impact of positive costimulation on both approaches was tested using our previously established Ba/F3 model system (Marcinek et al., 2023), which is devoid of any human costimulatory molecules. As a surrogate for synapse formation, conjugate formation as well as consecutive T-cell degranulation were studied. Further, we studied the cell death pathway mediating cytotoxicity against AML target cells, e.g. FAS/FASL vs perforin/granzyme B. The impact of continuous stimulation by BiTE®-and CART on evolving T-cell exhaustion was tested using our previously established 28-day long-term culture assay (Philipp et al., 2022). Finally, we compared BiTE® and CAR in an in vivo xenograft AML model. In a second in vivo experiment, all mice were harvested 18 days after T-cell injection, followed by T-cell isolation from the bone marrow and subsequent bulk RNA sequencing. BiTE®- and CAR-redirected T cells led to comparable effector-to-target (E:T) ratio-dependent specific lysis of various AML cell lines and pAML cells. Using our in vitro mixing experiments, we could show that BiTE®- and CAR-redirected T cells efficiently killed pAML cells while mostly sparing the hBM. Using the Ba/F3 model system, we observed that the expression of the costimulatory molecule CD86 significantly increased the cytotoxicity of BiTE®-redirected T cells while CAR-mediated lysis was unaltered. Furthermore, we were able to show that the percentage of T cell-AML conjugates formed as well as degranulation of CD107a was significantly higher when using CART compared to BiTE® molecules. Interestingly, when we added a FAS-blocking antibody to our cocultures, we could show that CART-mediated lysis of the target cells was significantly reduced. This was not observed when using the BiTE® construct. Using our long-term culture system, we found that T-cell proliferation and cytokine secretion decreased with both constructs over time. The cytotoxic capacity of BiTE®-redirected T cells seemed to decrease faster compared to the CART construct. Moving to an in vivo AML xenograft model, we observed that CART-treated mice had an improved OS compared to the BiTE®-treated mice (Fig.1). This was accompanied by enhanced T-cell proliferation and splenic homing of the CAR T cells. Bulk RNA sequencing of T cells isolated from the murine bone marrow 18 days post T-cell injection revealed signs of exhaustion in BiTE®-redirected T cells compared to CAR T cells. We found an upregulation of several inhibitory immune receptors and transcription factors (Fig.2) that have been well described to promote T-cell exhaustion. Furthermore, gene set enrichment analysis revealed that genes upregulated in effector vs exhausted were also upregulated in CAR T cells. This was accompanied by an upregulated glycolysis and fatty acid metabolism in CAR T cells. We conclude, that FLT3 is a promising target antigen with limited on-target off-leukemia toxicity. Our preclinical in vitro data demonstrate similar cytotoxicity of both platforms against AML cell lines and pAML cells. Utilizing a xenograft mouse model, CAR T cells provided a significant survival benefit over BiTE® molecules. Our data support the hypothesis that positive costimulation integrated within the CAR construct provided an advantage in T-cell activation and might lead to better T-cell homing, resulting in improved efficacy and delayed T-cell exhaustion in vivo. Future studies will be needed to further dissect differences between BiTE® vs CAR T cell-based immunotherapy platforms and identify suitable combination partners.

Single-Cell RNA Sequencing Reveals Contradictory Immune Microenvironment in Hemophagocytic Syndrome

Hemophagocytic lymphohistiocytosis (HLH), a formidable and rare clinical syndrome impacting multiple organs, manifests as a perilous condition. However, the intricate pathophysiological mechanisms underlying HLH remain yet to be fully elucidated. In this study, we obtained peripheral blood samples from 9 HLH patients spanning the years 2018 to 2022. Employing single-cell RNA sequencing (scRNA-seq), we conducted a comprehensive analysis of the alterations in the immune microenvironment, delving into the role of EBV infection in HLH and its influence on pathogenesis. By comparing with health donor peripheral blood , we identified a total of 11 significant immune cell populations among 72,779 single-cell transcriptomes, comprising 20,337 cells from healthy controls and 52,442 cells from HLH patients. Notably, we found that the proportion of T cells increased significantly, and other immune cells decreased in HLH patients. In terms of function, we found that the overall T cell killing ability was weakened, and the factor secretion ability was enhanced. Concerning myeloid cells, the impact of CD16+ monocytes on T cells exhibited a pronounced increase, along with an enhanced ability to secrete IL-18 factors, potentially serving as the main source of cells promoting T cell function. Different subpopulations of T cells have significant changes in the pathogenesis of HLH. Remarkably, both CD4+ T cells and CD8+ T cells exhibited HLA-DR+GZMK+ cell subgroups with upregulated IFN-γ production and response, suggesting their association with the hyperinflammatory state in HLH. A correlation analysis between the proportion of HLA-DRA+ GZMK+ T cells and the severity score revealed a significant correlation (p&amp;lt;0.05), with the ROC curve further affirming their predictability in assessing high inflammatory activity. Among CD4+ T cells, the proportion of Treg cells and the secretion of inhibitory factors such as TGFb1 decreased. Additionally, we noted the presence of a functionally impaired subset of CBLB+CD8+ T cells, indicating the considerable variability in T cell subset functionality in HLH. Next, a comprehensive analysis of the alterations in myeloid cells, B cells, and NK cells in HLH patients was conducted. We observed a substantial reduction in the number of DC cells and immune-related functions within various myeloid cell subsets. Regarding B cells, an increase in anergy B cells was observed, whereas the population of plasma cells, an effective B cell type, exhibited a decrease. In the context of NK cells, there was a significant increase in the proportion of NK cell subsets associated with IFN-γ secretion. Moreover, EBV-infected T cells in EBV-associated HLH were identified as the principal source of IFN-γ. Through single-cell EBV-CALL analysis, we found that IFN-γ secretion was primarily attributed to EBV-infected T cells, subsequently influencing CD16+ monocytes. Additionally, IL-16 production was predominantly observed in EBV-infected CD16+ monocytes, thereby impacting lymphocytes. Moreover, Monocytes were stimulated by EBV-infected CD4+ T cells and Atypical T cells, leading to the secretion of a pro-inflammatory factors such as MIF. Furthermore, our investigation unveiled that EBNA2 could promote MYC into the nucleus and enhance the proliferation and IFN-γ secretion of T cells. In conclusion, our comprehensive analysis of the immune microenvironment changes following the onset of HLH has yielded valuable insights and novel perspectives for understanding the underlying mechanisms and potential clinical treatments of this condition.

Screening Dolosigranulum pigrum Strains for Secreted Factors that InhibitStaphylococcus aureus

Abstract Background Staphylococcus aureus is a leading cause of community-associated and hospital-acquired infections, and methicillin-resistant S. aureus (MRSA) is a significant global health concern. Approximately 30% of the human population is asymptomatically colonized by S. aureus in the upper respiratory tract, with this colonization typically preceding respiratory infections caused by S. aureus. Dolosigranulum pigrum is a putatively beneficial bacterium that is a frequent colonizer of the human upper respiratory tract. The presence and abundance of D. pigrum in the upper respiratory tract has been negatively associated with S. aureus colonization among infants and young children. Additionally, in cocultivation experiments, D. pigrum has been shown to inhibit S. aureus growth, although the mechanisms for this antistaphylococcal activity have not been identified. Methods We hypothesized that D. pigrum strains secrete a protein factor that inhibits growth of S. aureus. To evaluate this hypothesis, we screened D. pigrum strains for antistaphylococcal activity using a plate-based cocultivation assay. Briefly, 5 µL aliquots of D. pigrum strains in liquid brain heart infusion (BHI) medium (OD600=0.3) were spotted onto a BHI agar plate and incubated at 37°C and 5% CO2 for 48 hours. Next, 5 µL aliquots of S. aureus strain Newman in liquid BHI medium (OD600=0.1) were spotted onto this plate at varying distances from the D. pigrum inocula and incubated at 37°C and 5% CO2 for an additional 24 hours. Secretion of an anti-staphylococcal factor was assessed based on the presence of a visible zone of inhibition between the D. pigrum and S. aureus inocula. We excised agar fragments adjacent to 4 inhibitory and 4 non-inhibitory D. pigrum strains and submitted these samples for liquid chromatography with tandem mass spectrometry (LC-MS/MS)-based proteomics. The proteomic data were searched against the TrEMBL D. pigrum database and a common contaminant/spiked protein database, and signal intensity values for identified peptides were quantified. Results Four of 43 (9%) D. pigrum strains screened in cocultivation assays consistently produced a zone of inhibition against S. aureus. We identified 531 unique proteins from proteomic analysis of agar fragments, including 28 D. pigrum proteins that were significantly more abundant in zones adjacent to inhibitory vs. non-inhibitory strains. The most differentially abundant protein was an S8 family serine protease (112 times more abundant in fragments adjacent to inhibitory strains). Using previously generated whole-genome sequencing data from these 43 D. pigrum strains, we found that the gene encoding this serine protease was present in 41 of 43 (95%) D. pigrum strains, including all 4 strains that were inhibitory in the cocultivation assay. The predicted active domain for the final protein was highly conserved across strains. Other secreted D. pigrum proteins that were more abundant in gel fragments adjacent to inhibitory strains included a peptidoglycan hydrolase and 7 uncharacterized D. pigrum proteins. Conclusion D. pigrum inhibits S. aureus growth in vitro through secretion of inhibitory factors in a strain-dependent manner. We identified an S8 family serine protease as a possible candidate mechanism for the anti-staphylococcal activity of D. pigrum. Future experiments will screen cell-free supernatants from inhibitory D. pigrum strains for S. aureus inhibition and evaluate the effect of protease inhibitor treatment on the antistaphylococcal activity of these supernatants.

Associated factors of diabetic retinopathy by artificial intelligence evaluation of fundus images in Japan

AbstractThis cross-sectional study aimed to investigate the promoting and inhibitory factors of diabetic retinopathy (DR) according to diabetes mellitus (DM) stage using standardized evaluation of fundus images by artificial intelligence (AI). A total of 30,167 participants underwent blood and fundus examinations at a health screening facility in Japan (2015–2016). Fundus photographs were screened by the AI software, RetCAD and DR scores (DRSs) were quantified. The presence of DR was determined by setting two cut-off values prioritizing sensitivity or specificity. DM was defined as four stages (no DM: DM0; advanced DM: DM3) based on treatment history and hemoglobin A1c (HbA1c) levels. Associated factors of DR were identified using logistic regression analysis. For cutoff values, multivariate analysis revealed age, sex, systolic blood pressure (SBP), smoking, urinary protein, and HbA1c level as positively associated with the risk of DR among all DM stages. In addition to glycemic control, SBP and Fibrosis-4 index might act as promoting factors for DR at all or an earlier DM stage. T-Bil, cholinesterase, and T-cho level might be protective factors at an advanced DM stage.

Neutrophil to lymphocyte ratio (NTLR) predicts local control and overall survival after stereotactic body radiotherapy (SBRT) in metastatic sarcoma

The neutrophil to lymphocyte ratio (NTLR) and absolute lymphocyte count (ALC) recovery are prognostic across many cancers. We investigated whether NLTR predicts SBRT success or survival in a metastatic sarcoma cohort treated with SBRT from 2014 and 2020 (N = 42). Wilcox Signed Rank Test and Friedman Test compare NTLR changes with local failure vs. local control (N = 138 lesions). Cox analyses identified factors associated with overall survival. If local control was successful, NLTR change was not significant (p = 0.30). However, NLTR significantly changed in patients with local failure (p = 0.027). The multivariable Cox model demonstrated higher NLTR before SBRT was associated with worse overall survival (p = 0.002). The optimal NTLR cut point was 5 (Youden index: 0.418). One-year overall survival in SBRT metastatic sarcoma cohort was 47.6% (CI 34.3%–66.1%). Patients with an NTLR above 5 had a one-year overall survival of 37.7% (21.4%–66.3%); patients with an NTLR below 5 had a significantly improved overall survival of 63% (43.3%–91.6%, p = 0.014). Since NTLR at the time of SBRT was significantly associated with local control success and overall survival in metastatic sarcoma treated with SBRT, future efforts to reduce tumor inhibitory microenvironment factors and improve lymphocyte recovery should be investigated.

PENERAPAN KEBIJAKAN E-GOVERNMENT SISTEM APLIKASI E-KINERJA BERBASIS WEB DI LINGKUNGAN PEMERINTAH KABUPATEN MUKOMUKO

The purpose of this study is to find out how E-Government policy issues through the web-based E-performance application system in the Mukomuko Regency government environment. This research uses qualitative descriptive research and data collection through observation, interviews, and documentation, with the intention of being able to objectively describe facts and phenomena that are research problems. The results showed that the issue of E-Government policy through the implementation of a web-based E-performance application system in the Mukomuko Regency government environment has not been going well. This is because there are still shortcomings in the E-performance application system used and several inhibitory factors found. The limitations of research specifically only address e-government policy issues through the implementation of a web-based E-performance application system in mukomuko regency. This research can be used as a reference for innovation and policy development in the field of E-government through the concept of innovation adaptation.

Examination of the Factors Inhibiting CO2 Adsorption on Coal: A Case Study from Shallow-Depth Low-Rank Coal Seams.

Understanding the inhibitory factors affecting the adsorption of CO2 on low-rank coal from shallow-depth coal seams is essential to identify potential coal seams for CO2 sequestration. The CO2 adsorption capacity of shallow-depth coals was measured at a low pressure on raw and dry coals. The samples were also prepared for organic analyses, scanning electron microscopy analyses, and low-temperature nitrogen adsorption analyses to evaluate the CO2 adsorption and identify the inhibitory factors. An investigation was conducted to determine how CO2 adsorption occurs on coal by fitting experimental data to adsorption isotherm models, followed by analyzing the results based on the statistical analysis. In addition, this study used Henry's law, surface potential, and Gibbs free energy to identify the adsorption inhibitor between CO2 and coal. The CO2 adsorption experiment was conducted on raw coal with a moisture content of 15.18-20.11% and dry coal with no moisture. The experimental data showed that the CO2 adsorption capacity in dry coal was 1.6-1.8 times greater than that in raw coal. A fitting graph between the adsorption data and the isotherm model indicated that CO2 adsorption on coal occurred on monolayers and multilayers under raw and dry conditions. Statistical evaluation of the adsorption isotherm models showed that the Langmuir and Freundlich models aligned more closely to the experimental data. According to this result, low-pressure adsorption of CO2 on coal occurred in monolayers and multilayers under raw and dry conditions. Coal containing a high huminite content had a higher potential for CO2 adsorption, and the drying increased the positive relationship. On the other hand, coal containing high inertinite content inhibited CO2 adsorption onto the coal, but the drying process did not adversely affect CO2 adsorption. Furthermore, coal with high moisture and inertinite content inhibited the affinity, accommodation, and spontaneous CO2 adsorption onto the coal. CO2 adsorption could lead to swelling, but moisture loss opened more sites and micropores, resulting in the swelling effect not closing all micropores in dry coal. Based on these results, coal seams with low moisture and inertinite content are the most promising for CO2 adsorption. Altogether, this study provides an understanding of the percentage of inhibitor factors that affects CO2 adsorption on low-rank coal from shallow depths, which may lead to different CO2 adsorption capacities.

Identifying factors that affect the use of health information technology in the treatment and management of hypertension

BackgroundWe conducted this study with the aim of identifying factors that affect the use of health information technology in the treatment and management of hypertension.MethodsThis paper is a descriptive-analytic study conducted in 2022. To obtain relevant articles, databases including Scopus, Web of Science, IEEE, and PubMed were searched and the time period was between 2013 and 2022. Based on the review of similar articles, a five-point Likert scale checklist was developed in the second phase. The statistical population of the present study was specialist physicians (N = 40) and patients (N = 384). In order to analyze the data, SPSS Statistics 24 was used. To analyze the data obtained from the checklist, we used summary statistics (mean and standard deviation).ResultsAs a result of the review literature process, 50 papers were screened, that based we can distinguish motivational and inhibitory factors affecting the use of health information technology in hypertension management. Indeed, Motivational factors and inhibitory factors can be classified into five groups: organizational, economic, technical, personal, and legal/moral factors. Based on the results of the checklist, the factors that were identified as most influential on motivation and inhibitory patients and specialist physicians’ to use of health information technology to manage and treat hypertension.ConclusionUtilizing technologies for hypertension, its management can be improved by identifying motivating and inhibiting factors. Our approach can improve the acceptability of these technologies, save costs, reduce long-term complications of hypertension, and improve patient quality of life.

Senescent Schwann cells induced by aging and chronic denervation impair axonal regeneration following peripheral nerve injury

Following peripheral nerve injury, successful axonal growth and functional recovery require Schwann cell (SC) reprogramming into a reparative phenotype, a process dependent upon c‐Jun transcription factor activation. Unfortunately, axonal regeneration is greatly impaired in aged organisms and following chronic denervation, which can lead to poor clinical outcomes. While diminished c‐Jun expression in SCs has been associated with regenerative failure, it is unclear whether the inability to maintain a repair state is associated with the transition into an axonal growth inhibition phenotype. We here find that reparative SCs transition into a senescent phenotype, characterized by diminished c‐Jun expression and secretion of inhibitory factors for axonal regeneration in aging and chronic denervation. In both conditions, the elimination of senescent SCs by systemic senolytic drug treatment or genetic targeting improved nerve regeneration and functional recovery, increased c‐Jun expression and decreased nerve inflammation. This work provides the first characterization of senescent SCs and their influence on axonal regeneration in aging and chronic denervation, opening new avenues for enhancing regeneration and functional recovery after peripheral nerve injuries.

Differentiated Instruction in Indonesian Private Kindergartens: Challenges in Implementing an Independent Curriculum

Differentiated instruction facilitates learning based on students’ unique interests, preparedness for learning, and learning profiles. Many teachers lack a comprehensive understanding of differentiated instruction, particularly in kindergarten. Teachers used to employ Student Worksheets to facilitate learning. This study aims to: (1) the implementation of differentiated instruction at an Indonesian private kindergarten, (2) the factors that facilitate the instruction’s implementation, and (3) the efforts to deal with any challenges encountered during the implementation of the instruction. This study employed a qualitative case study design to capture a phenomenon comprehensively and contextually by gathering data directly from the subject under investigation. The study was executed at Aisyiyah Kindergarten, Sukorejo, Indonesia. It is the only Indonesian kindergarten in Sukorejo that incorporates an independent curriculum called Sekolah Penggerak. Teachers implement holistic-integrative learning design to differentiate instructional activities. Learning media containing STEAM Loose parts were utilized to determine various factors that may facilitate or impede the instruction. Factors that facilitate instruction include the learning environment, media utilization, teachers’ creativity, the student-teacher relationship, the implementation of teaching techniques, students’ psychological well-being, and community support beyond the school setting. The inhibitory factors encompass a scarcity of learning resources, students’ lack of preparedness for learning, a shortage of teachers, and inadequate infrastructure. This study revealed that the school tried to address the challenges by collaborating with the community inside and outside the school, such as with student guardians, the foundation administrator, and related agencies